Objective To evaluate the effect of internal fixation on the stability of pedicled fascial flap and the osteogenesis of exceed critical size defect (ECSD) of bone so as to provide theory for the clinical application by the radiography and histology observation. Methods The ECSD model of the right ulnar midshaft bone and periosteum defect of 1 cm in length was established in 32 New Zealand white rabbits (aged 4-5 months), which were divided into group A and group B randomly (16 rabbits in each group). The composite tissue engineered bone was prepared by seeding autologous red bone marrow (ARBM) on osteoinductive absorbing material (OAM) containing bone morphogenetic protein and was used repair bone defect. A pedicled fascial flap being close to the bone defect area was prepared to wrap the bone defect in group A (control group). Titanium miniplate internal fixation was used after defect was repair with composite tissue engineered bone and pedicled fascial flap in group B (experimental group). At 2, 4, 6, and 8 weeks, the X-ray films examination, morphology observation, and histology examination were performed; and the imaging 4-score scoring method and the bone morphometry analysis was carried out. Results All rabbits survived at the end of experiment. By X-ray film observation, group B was superior to group A in the bone texture, the space between the bone ends, the radiographic changes of material absorption and degradation, osteogenesis, diaphysis structure formation, medullary cavity recanalization. The radiographic scores of group B were significantly higher than those of group A at different time points after operation (P lt; 0.05). By morphology and histology observation, group B was superior to group A in fascial flap stability, tissue engineered bone absorption and substitution rate, external callus formation, the quantity and distribution area of new cartilage cells and mature bone cells, and bone formation such as bone trabecula construction, mature lamellar bone formation, and marrow cavity recanalization. The quantitative ratio of bone morphometry analysis in the repair area of group B were significantly larger than those of group A at different time points after operation (P lt; 0.05). Conclusion The stability of the membrane structure and the bone defect area can be improved after the internal fixation, which can accelerate bone regeneration rate of the tissue engineered bone, shorten period of bone defect repair, and improve the bone quality.